Fibrin will be the normal scaffold for cells to invade in tissue repair. For several years, however, natural or synthetic polymers have been used for forming artificial scaffolds being applied to the damaged tissue for improving the repair process. Tissues are generally characterized in respect to functionality and appearance and are classified as hard or soft tissue. Soft tissue can further be categorized in cartilage and tendons opposed to loose connective tissue and muscles. The invention relates to both soft and hard tissue and the interface between these like in respect to cartilage and bone. Further the invention relates to synthetic scaffolds and to dermatan sulfate and derivatives of dermatan sulfate.
Cartilage is dense tissue composed of collagenous fibers and/or elastic fibers formed by the growth of chondrocytes all embedded in a firm gel-like matrix.
The matrix is mainly composed of proteoglycans filling the space between collagen fibers and is able to hold water. Continued synthesis, assembly and degradation by the chondrocytes maintain the structure and structural integrity of the extracellular matrix (ECM) of cartilage.
The proteoglycans are composed of a protein backbone and glycosamino glycans (GAG) linked to this. In the articular cartilage, the proteoglycans are of the aggrecan type containing mainly chondroitin- and keratan sulfate. Besides maintaining the physical and structural integrity of the cartilage, the GAGs are also involved in the cell adhesion, migration, proliferation and differentiation (Plaas A H K, Wong-Palms S, Roughley P J, Midure R J & Hascall V C. Chemical and immunological assay of the nonreducing terminal residues of chondroitin sulfate from human aggrecan. J. Biol. Chem. 272, 20603-20610 (1997)). Furthermore the GAGs form a network that protects the chondrocytes from the potentially damaging forces of mechanical function.
The function of the cartilage includes providing a framework upon which bone deposition can begin and supply a smooth surface for the movement of articulating bone. Cartilage is found in many different places in the body and can be found in three different types: hyaline, elastic and fibrocartilage. The different forms have special characteristics adapted to their function with the hyaline form being the most abundant type of cartilage. This form, named articular cartilage, is predominantly made of type II collagen and found lining bones in joints. This form of cartilage provides the joint with a low friction and wear-resistant bearing surface, that can tolerate a tremendous amount of repetitive physical stress.
Damaged articular cartilage has very little capacity for spontaneous healing, because of the hypocellularity and absence of both vascularization and innervation in the cartilage to support repair and remodeling.
Because of this lacking capacity for self-healing, many attempts have been made to facilitate the healing using scaffolds. As an attempt to mimic the ECM of cartilage, these are often made by incorporating various GAGs in a matrix of either natural or synthetic polymer. Important classes of GAGs are Hyaluronic acid (HA), Chondroitin Sulphate (CS), Heparan Sulfate (HS), Heparin, Keratan sulfate (KS) and Dermatan Sulphate (DS).
Yen-Lin Chen et al. (Composite chondroitin-6-sulfate/dermatan sulfate/chitosan scaffolds for cartilage tissue engineering. Biomaterials (2007)) describes the use of chitosan as the base material. A solution of this is freeze-dried to porous scaffolds, and varying amounts of CS and DS are bound covalently to this matrix. These scaffolds were seeded with chondrocytes. Overtly higher expression levels of aggrecan and collagen II was found in the CS+DS-containing scaffolds compared to the DS-only scaffolds. Cells in the DS-only scaffolds were clustered and no apparent lacuna was observed. DS-only was found to enhance GAG and collagen production, but did not stimulate cell proliferation.
Chic-Ta Lee, Ching-Ping Huang & Yu-Der Lee. (Biomimetic porous scaffolds made from poly(L-lactide)-g-chondroitin sulfate blend with poly(L-lactide) for cartilage tissue engineering. Biomacromolecules 7, 2200-2209 (2006)) describes how PLA is grafted to CS and a composite of this and PLA is made into porous scaffolds by solvent casting/particle leaching. These are seeded with mouse chondrocytes and cell adhesion, secretion of ECM, the quantity of synthesized collagen and GAGs and the compression modulus was then examined. The scaffold stimulated the growth of new cartilage, and after 4 weeks, the compression modulus was close to that of mouse cartilage.
Hyuk Sang Yoo, Eun Ah Lee, Jun Jin Yoon & Tae Gwan Park. (Hyaluronic acid modified biodegradable scaffolds for cartilage tissue engineering. Biomaterials 26, 1925-1933 (2005)) describes a blend of PLGA and PLGA-PEG-NH2 diblock copolymer made into porous scaffolds, and then HA is grafted to the surface. This is seeded with chondrocytes. It is concluded that the HA-immobilized scaffolds help the chondrocytes to retain their phenotype to a greater extent than the unmodified scaffold, and growth of new cartilage is observed within one month.
In Hongbin Fan et al. (Cartilage regeneration using mesenchymal stem cells and a PLGA-gelatin/chondroitin/hyaluronate hybrid scaffold. Biomaterials 26, 4573-4580 (2006)) a composite scaffold of PLGA and cross-linked gelatin/CS/HA is made and seeded with mesenchymal stem cells. The cells are induced to differentiate to chondrocytes, and seeded onto the scaffolds. In-vitro proliferation and GAG synthesis is examined. Scaffolds (both with and without gelatin/CS/HA) seeded with cells are implanted in rabbits with cartilage defects, and the rabbits are harvested after 6, 12 and 24 weeks. Both in-vitro and in-vivo results show superior results for the GAG-modified scaffold compared to plain PLGA. Both types gave formation of new cartilage in the rabbits, but the plain PLGA gave a thinner cartilage with inferior morphology.
Job L. C. van Susante et al. (Linkage of chondroitin-sulfate to type I collagen scaffolds stimulates the bioactivity of seeded chondrocytes in vitro. Biomaterials 22, 2359-2369 (2007)) describes how a solution of type I collagen is freeze dried to porous scaffolds. CS is then grafted to these and the scaffolds are seeded with chondrocytes. Cell proliferation and was higher in the CS-modified scaffold and growth of cartilage was better than in unmodified scaffolds.
In all the examples above, GAGs are grafted covalently to the base material. Because both CS and HA are abundant in the ECM of cartilage, these are often the first choice when making scaffolds.